Title: LC-MS/MS as an alternative for SDS-PAGE in blue native analysis of protein complexes
Authors: Wessels, Hans J C T
Vogel, Rutger O
van den Heuvel, Lambertus
Smeitink, Jan A
Rodenburg, Richard J
Nijtmans, Leo G
Farhoud, Murtada H # ×
Issue Date: Sep-2009
Publisher: WILEY-VCH Verlag
Series Title: Proteomics vol:9 issue:17 pages:4221-8
Abstract: Two-dimensional blue native/SDS-PAGE is widely applied to investigate native protein-protein interactions, particularly those within membrane multi-protein complexes. MS has enabled the application of this approach at the proteome scale, typically by analysis of picked protein spots. Here, we investigated the potential of using LC-MS/MS as an alternative for SDS-PAGE in blue native (BN) analysis of protein complexes. By subjecting equal slices from BN gel lanes to label-free semi-quantitative LC-MS/MS, we determined an abundance profile for each protein across the BN gel, and used these profiles to identify potentially interacting proteins by protein correlation profiling. We demonstrate the feasibility of this approach by considering the oxidative phosphorylation complexes I-V in the native human embryonic kidney 293 mitochondrial fraction, showing that the method is capable of detecting both the fully assembled complexes as well as assembly/turnover intermediates of complex I (NADH:ubiquinone oxidoreductase). Using protein correlation profiling with a profile for subunits NDUFS2, 3, 7 and 8 we identified multiple proteins possibly involved in the biogenesis of complex I, including the recently implicated chaperone C6ORF66 and a novel candidate, C3ORF60.
ISSN: 1615-9853
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Non-KU Leuven Association publications
× corresponding author
# (joint) last author

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